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L K Trejdosiewicz

Publications and source records attributed to L K Trejdosiewicz.

At least 37 records · Page 2Linked to original sources
PubMed1999

Cytokeratin expression patterns in normal and malignant urothelium: a review of the biological and diagnostic implications.

The cytokeratins are the intermediate filament proteins characteristic of epithelial cells. In human cells, some 20 different cytokeratin isotypes have been identified. Epithelial cells express between two and ten cytokeratin isotypes and the consequent profile which reflects both epithelial type and differentiation status may be useful in tumour diagnosis. The transitional epithelium or urothelium of the urinary tract shows alterations in the expression and configuration of cytokeratin isotypes related to stratification and differentiation. In transitional cell carcinoma, changes in cytokeratin profile may provide information of potential diagnostic and prognostic significance. The intensification of immunolabelling with some CK8 and CK18 antibodies may underly an active role in tumour invasion and foci of CK17-positive cells may represent proliferating populations. Loss of CK13 is a marker of grade and stage and de novo expression of CK14 is indicative of squamous differentiation and an unfavourable prognosis. However, perhaps the most important recent finding is the demonstration that a normal CK20 expression pattern is predictive of tumour non-recurrence and can be used to make an objective differential diagnosis between transitional cell papilloma and carcinoma. This review will consider cytokeratin expression in urothelium and discuss the application of cytokeratin typing to the diagnosis and prognosis of patients with TCC.

Biomarkers↗
PubMed1998

Interleukins 4 and 13 upregulate expression of cd44 in human colonic epithelial cell lines.

Interleukin 4 (IL-4) inhibits carcinoma cell growth and promotes expression of differentiation-associated products by normal and malignant epithelial cells. The effects of IL-4 and IL-13 on expression of the CD44 transmembrane adhesion receptor were examined in human epithelial cell lines of colonic (HT-29, CaCo-2, DLD-1, T84), breast (MCF-7, ZR75-1) and liver (Hep-G2, PLC/PRF/5) origins as well as mitogen-activated and resting peripheral blood lymphocytes (PBL) and T cell lines (Jurkat, HUT78). Liver and Jurkat cells were negative for CD44. Colonic, breast and HUT78 cells expressed CD44 constitutively and all except DLD-1 and HUT78 also expressed CD44 splice variant (CD44v) epitopes. All cell lines expressed IL-4 receptors, but IL-4 and IL-13 induced upregulation of CD44 only in the colonic cell lines. CD44v was also upregulated, but there was no de novo induction of CD44v in variant-negative cells and no de novo expression of CD44 in the CD44(-) lines. CD44 upregulation in mitogen-activated PBL was not increased by IL-4 and IL-13 and was not inhibited by neutralizing antibodies. Other cytokines tested [interferon gamma (IFN-gamma, tumour necrosis factor alpha (TNF-alpha), transforming growth factor beta1 (TGF-beta1) and IL-6] did not affect CD44 core epitope expression in the cell lines tested.

Alternative Splicing↗
PubMed1998

Uroplakin gene expression by normal and neoplastic human urothelium.

cDNA sequences for human uroplakins UPIa, UPIb, UPII, and UPIII were cloned and used to investigate uroplakin transcription by normal and neoplastic urothelial cells. Normal urothelium expressed mRNA for all four uroplakins, although UPIII could be detected only by ribonuclease protection assay. By in situ hybridization, UPIa and UPII were confined to superficial cells and UPIb was also expressed by intermediate cells. Cultured normal human urothelial cells showed a proliferative basal/intermediate cell phenotype and constitutive expression of UPIb only. Uroplakin expression by transitional cell carcinoma cell lines was related to their differentiated phenotype in vitro. RT4 cells expressed all uroplakins, VM-CUB-3 expressed three uroplakins, RT112 and HT1376 cells expressed only UPIb in high abundance, and COLO232, KK47, and EJ cells had no detectable expression. These results correlated with patterns of uroplakin expression in tumors. UPIa and UPII were detected superficially only in well differentiated transitional cell carcinoma papillae. UPIb was positive in seven of nine and overexpressed in five of nine noninvasive transitional cell carcinomas and was also present in four of eight invasive transitional cell carcinomas. Lymph node metastases retained the same pattern of UPIb expression as the primary tumor. Unlike the three differentiation-regulated uroplakins, UPIb may have an alternative role in urothelial cell/tissue processes.

Amino Acid Sequence↗
PubMed1998

Expression and cytokine regulation of immune recognition elements by normal human biliary epithelial and established liver cell lines in vitro.

BACKGROUND/AIMS: Biliary epithelial cells are targets of immune-mediated attack in conditions such as primary biliary cirrhosis and allograft rejection. This has been attributed to the ability of biliary epithelial cells to express ligands for T cell receptors. We aimed to investigate the expression of immune recognition elements and the effects of pro-inflammatory and anti-inflammatory cytokines on cell surface phenotypes of normal human biliary epithelial cells and established human liver-derived (PLC/PRF/5, HepG2, Hep3B and CC-SW) lines. METHODS: Cells were cultured in the presence or absence of cytokines for 72 h, and expression of cell surface molecules was assessed by flow cytometry and immunofluorescence. RESULTS: All cell lines expressed MHC class I, ICAM-1 (CD54), LFA-3 (CD58) and EGF receptor, and all but Hep3B expressed Fas/Apo-1 (CD95). Unlike hepatocyte-derived cell lines, biliary epithelial cells and CC-SW expressed CD40 and CD44. As expected, IFNgamma and TNFalpha upregulated expression of ICAM-1, MHC class I and MHC class II, particularly in biliary epithelial cells. TGFbeta downregulated these molecules and downregulated CD95 on biliary epithelial cells, but upregulated LFA-3. The Th2 cytokines had little effect, although IL-4 upregulated CD95 expression on biliary epithelial cells. IFNgamma upregulated CD40 expression on biliary epithelial cells, CC-SW and HepG2. CONCLUSIONS: These findings imply that biliary epithelial cells may be capable of interacting with activated T lymphocytes via CD40 and LFA-3, which are thought to be important T cell accessory ligands for T cell activation in a B7-independent manner. Sensitivity to pro-inflammatory cytokines and expression of CD95 may explain why biliary epithelial cells are primary targets for autoimmune attack.

Bile Ducts, Intrahepatic↗
PubMed1998

Effects of resuscitation fluids on T cell immune responses.

This study was designed to determine whether dextran, gelatin or hydroxyethyl starch-based colloidal resuscitation fluids (CRF) are inhibitory to T lymphocyte activation and mitogenesis in vitro. Isolated peripheral blood lymphocytes from normal donors were activated with mitogen (PHA) and cultured in up to 50% v:v CRF. Dual-label flow cytometry for CD69 and CD25 were used to assess early and full T cell activation responses, respectively, and thymidine incorporation was used to assess mitogenesis. T cell activation and mitogenic responses were not inhibited in the presence of CRF, implying that any systemic immunodepression associated with CRF infusion is not directly related to CRF-mediated impairment of T cell activation.

Adult↗
PubMed1998

Culture of ascitic ovarian cancer cells as a clinically-relevant ex vivo model for the assessment of biological therapies.

There are few in vitro models of human ovarian cancer suitable for the assessment of biological therapies. We have established short-term cultures of ovarian carcinoma cells from ascites, maintained in suspension in ascitic fluid to help preserve the original tumor cell microenvironment. We assessed the effects of a potential biological therapeutic agent, interferon-alpha (IFN alpha), on ovarian cell phenotype. In total, eight cultures were established from seven patients. Quantitative changes in cell surface phenotype were determined by flow cytometry for HLA-ABC, HLA-DR, TAG72, CA125, HMFG1 and HMFG2 antigens. The amount of CA125 antigen shed into the culture media was also assessed. Variations in cell surface phenotype between specimens probably reflected the specific cytokine milieu of the ascites as well as idiotypic differences between tumors. Nevertheless, there were consistent phenotypic responses to IFN alpha, with up-regulation of MHC Class I but down-regulation of the HMFG1 and HMFG2 antigens from the cell surface. The results suggest that this approach may be useful in patient selection and for optimizing biological therapies, as it enables patients' individual tumor responses to exogenous cytokine to be studied against the background of the endogenous cytokine milieu.

Aged↗
PubMed1997

Effects of resuscitation fluids on nonadaptive immune responses.

BACKGROUND: Colloidal plasma-expander fluids are commonly used as an alternative to blood components in the resuscitation of patients suffering from hemorrhagic shock and trauma. Of these, hydroxyethyl starch is also used as a cryopreservative, and these dual properties have been utilized in the development of a blood storage system that allows the direct transfusion of red cells. The prolonged intravascular persistence of hydroxyethyl starch suggests that phagocytic clearance may be impaired and that the presence of hydroxyethyl starch could exacerbate transfusion-induced immunomodulation. STUDY DESIGN AND METHODS: The effects of colloidal resuscitation fluids on the activation response and phagocytic function of polymorphonuclear cells (PMNs) and monocytes in normal peripheral blood were examined. To mimic the hemolysis associated with cryopreservation, the effects of 1- and 5- percent red cell lysate were studied. Flow cytometric assays were used in all cases. RESULTS: The percentage of phagocytic monocytes and PMNs was not altered; nor were the rates of phagocytosis impaired after incubation with resuscitation fluids. Upregulation of cell surface integrin during activation was similarly unmodified by the fluids. CONCLUSION: Hydroxyethyl starch and other resuscitation fluids do not affect some important antimicrobial functions of the nonadaptive arm of the immune response. This suggests that posttrauma or transfusion-induced immunomodulation is not exacerbated by inhibition at this level.

Adaptation, Physiological↗
PubMed1997

Reconstitution of human urothelium from monolayer cultures.

PURPOSE: We established a 3-dimensional organ culture model of urinary tract tissue in which to study the effects of seeding cultured urothelial cells onto de-epithelialized urothelial stroma. MATERIALS AND METHODS: Normal human urinary tract tissues were placed in organ culture or used to establish urothelial cell cultures. At passage 2 cell cultures were harvested and used to reconstitute autologous organ cultures by seeding onto de-epithelialized stroma. Organ cultures were harvested at intervals and analyzed by immunohistology with a panel of antibodies against differentiation associated antigens, cytokeratins, cell adhesion molecules, extracellular matrix components and proliferation associated antigens. RESULTS: Human urothelial tissues were maintained in organ culture for at least 18 weeks and they retained a transitional epithelial morphology with expression of normal in situ antigenic characteristics. Within 2 weeks of reconstitution recombined organ cultures formed a stratified, polarized, transitional-like neo-epithelium that expressed many of the phenotypic and differentiated characteristics of normal tissue. Basement membrane formed at sites of direct contact between urothelial cells and stroma. After an initial stabilization period the proliferation rate of the urothelium of intact and reconstituted organ cultures decreased to the low turnover rate characteristic of normal urothelium in situ, indicating that the cells were responsive to normal growth regulatory controls. CONCLUSIONS: Normal human urothelial cells, which express a proliferative nondifferentiated phenotype in monolayer culture, retain the capacity to differentiate and reform a slow turnover, stratified transitional epithelium.

Cell Adhesion↗
PubMed1997

Stromal and vascular invasion in an human in vitro bladder cancer model.

In transitional cell carcinoma (TCC) of the urinary bladder, disease spread may occur through local invasion of the bladder wall and/or via hematogenous dissemination. Although the metastatic phenotype is well studied, little is known about the mechanisms that determine and regulate secondary dispersion via stromal and vascular routes. The aim of this study was to develop an in vitro system to study TCC invasion using normal human urinary tract stroma in organ culture. Human de-epithelialized urinary tract stromas were seeded with established human TCC cell lines (RT4, RT112, and EJ) and maintained in organ culture at an air-liquid interface for up to 8 weeks. The composite tissues were analyzed immunohistochemically using antibodies to a proliferation-associated antigen, cell adhesion molecules, extracellular matrix components, and cytokeratin isotypes. The phenotype of TCC cell lines maintained as monolayer cultures were compared with the composite tissues to assess the regulatory effects of the stroma. On a normal urothelial stroma, the TCC cell lines proliferated and underwent a degree of differentiation related to the grade of the original tumor. RT4 cells formed a stratified epithelium with low proliferation, some characteristics of urothelial differentiation, and no evidence of stromal invasion over the 8-week culture period. RT12 cells formed a stratified epithelium with limited differentiation and by 2 weeks showed intravasation of the subepithelial capillary bed. Anaplastic EJ cells did not stratify but diffusely invaded the stromal matrix and the superficial vasculature. In conclusion, the urothelial stroma can support cytodifferentiation in vitro and influence the behavior of malignant TCC cells. The model has application for determining genetic and epigenetic factors that influence whether a tumor progresses by local and/or hematogenous spread. Furthermore, it may offer a useful approach in evaluating prognostic and diagnostic markers and strategies for preventing invasion.

Antibodies, Monoclonal↗
PubMed1996

The effects of dietary fatty acids on the proliferation of normal human urothelial cells in vitro.

Little is known of the mechanisms by which dietary fatty acids (FAs) may affect normal epithelial cell physiology and thereby directly or indirectly influence tumour incidence and progression. In this study, we have used normal human urothelial cell cultures to investigate whether FAs may modify proliferation of normal human epithelial cells in vitro. FAs were presented as albumin complexes in serum-free medium and the effects on proliferation over a concentration range of 1-100 microM were assayed by thymidine incorporation. Saturated FAs had no effect. At lower concentrations (1-10 microM), mono-unsaturated FAs (MUFAs) and n-3 polyunsaturated FAs (PUFAs) were slightly stimulatory. Concentrations of unsaturated FAs above 10 microM were growth inhibitory in a dose-dependent manner. Oleic acid showed least cytostatic effect, whereas gamma-linolenic acid induced irreversible growth arrest. Although marked morphological effects were observed in irreversibly growth-inhibited cells, the cells remained attached to the substratum and showed no evidence of nuclear pyknosis or apoptosis. The inhibitory effects of single PUFAs could be reduced, but not abolished, by the addition of saturated FAs or MUFAs. Mixtures of different PUFAs were inhibitory in an additive manner. These data suggest that PUFAs have a cytostatic effect on rapidly proliferating epithelial cells which appears unrelated to malignant transformation.

Analysis of Variance↗
PubMed1995

Patterns of splice variant CD44 expression by normal human urothelium in situ and in vitro and by bladder-carcinoma cell lines.

CD44 core and splice variant exon expression was investigated in the stratified transitional epithelium of the urinary tract and in normal and malignant bladder epithelial cell lines in vitro. Antibodies against core CD44 epitopes and splice exon variants v3, v4/5, v5 and v6 showed an intense reaction in the basal and lower intermediate urothelial cell layers, which was consistently lost from the upper intermediate and superficial cell layers. Seven independent cell lines established from normal human urothelium expressed complex multiple-spliced CD44 mRNA transcripts when tested by RT-PCR and were positive with antibodies against CD44 core epitopes and splice variants v3, v4/5, v5 and v6. Of the 13 bladder-carcinoma cell lines, all were positive for CD44 core. The more differentiated cell lines had retained some splice-variant antibody reactivity and showed multiple but less complex CD44 mRNA transcript patterns, compared with normal cells. Anaplastic cell lines did not react with variant antibodies and did not contain multiple-spliced CD44 transcripts. These data suggest that loss of alternatively-spliced CD44 may reflect a selection pressure in the evolution of anaplastic bladder cancers.

Antigens, Neoplasm↗
PubMed1995

T-cell responses and cellular immunity in coeliac disease.

Increasing evidence points to a direct role for T cells in the mediation of the coeliac intestinal lesion. There is good evidence for increased local T-cell reactivity, manifest as increased in T-cell activation in the lamina propria and T-cell proliferation in the epithelial compartment. A likely scenario is that gluten elicits antigen-specific responses by lamina propria T helper cells, probably of the Th1 (inflammatory-mediator) subtype, leading to secretion of pro-inflammatory cytokines. Such cytokines may have direct effects on intestinal enterocytes, as well as mediating indirect effects by upregulation of MHC antigens and by enhancing the activity of cytolytic T cells. Although gluten-specific IEL responses have not been demonstrated by intraepithelial T lymphocytes (IELs), increasing evidence suggests that IELs can act as cytolytic effector cells and hence are likely to exert enteropathic effects under the influence of pro-inflammatory cytokines.

Celiac Disease↗
PubMed1995

Peripheral cell-mediated immune response to mycobacterial antigens in inflammatory bowel disease.

A mycobacterial etiology has been proposed in Crohn's disease (CD). We have sought evidence of increased or modified T lymphocyte immune responses to Mycobacterium tuberculosis and Myco, paratuberculosis in patients with CD (n = 13), compared with ulcerative colitis (UC; n = 17) and controls (n = 17). Peripheral blood cells were cultured with phytohaemagglutinin (positive mitogen control), mycobacterial purified protein derivative (PPD) preparations, lysates, column fractions and whole, heat-killed bacteria. Responses of T cells and T cell subsets were assessed by expression of activation markers (CD25, CD69), coupled with blastogenesis assays (3H-thymidine uptake) and estimates of proliferation. Virtually all patients responded to Myco. paratuberculosis and Myco. tuberculosis antigens. There were no significant differences between patient groups, although there was a very high overall correlation (r = 0.95; P < 0.0001) between responses to the two mycobacterial species. Most of the activation and proliferative responses resided in the CD4+ (T helper) subset. Although up to 15% of CD8+ (suppressor/cytotoxic) cells also became activated, the CD8+ cells did not proliferate subsequently. Cells expressing the alternate gamma delta form of the T cell receptor (TCR gamma delta+) did not activate or proliferate in response to mycobacterial antigens. There were no differences in any of these parameters between patient groups. We conclude that there is no specific increase or alteration in cell-mediated anti-mycobacterial immunity in inflammatory bowel disease (IBD). Thus our data do not support a mycobacterial etiopathology of Crohn's disease.

Adult↗
PubMed1995

Mycobacterium paratuberculosis DNA not detected in Crohn's disease tissue by fluorescent polymerase chain reaction.

The role of mycobacteria in the aetiology of Crohn's disease has been a contentious subject for many years. Mycobacterium paratuberculosis is known to cause a chronic granulomatous enteritis in animals (Johne's disease) and has been implicated as a possible infectious cause of Crohn's disease. However this fastidious organism is only rarely detected by conventional microbiological techniques. This study used oligonucleotide primers to the species-specific M paratuberculosis IS900 DNA insertion element and the polymerase chain reaction to amplify any M paratuberculosis DNA from intestinal tissue DNA extracts. One oligonucleotide primer was fluorochrome-labelled and the presence of fluorescent amplified product was determined using an automated DNA sequencer with a computerised gel-scanning laser. This method was shown capable of detecting 1-2 mycobacterial genomes. Intestinal tissue samples were obtained from 68 patients with histologically confirmed Crohn's disease, 49 patients with histologically confirmed ulcerative colitis, and 26 non-inflammatory bowel disease controls. In no case was M paratuberculosis detected in any of the inflammatory bowel disease tissue samples and only one non-inflammatory bowel disease case was positive. These results do not support the hypothesis that M paratuberculosis has an aetiological role in Crohn's disease.

Adolescent↗
PubMed1995

Expression and in vitro regulation of integrins by normal human urothelial cells.

Integrins are thought to be essential adhesion receptors for the maintenance of tissue histioarchitecture. The purpose of this study was to determine integrin expression patterns in the human stratified transitional epithelium of the urinary tract (urothelium). In situ expression patterns were compared with in vitro expression, using a normal cell culture model system in which the effects of cell stratification can be studied independently of differentiation. By immunohistological criteria, the urothelia of bladder, ureter and renal pelvis expressed alpha 2 beta 1 and alpha 3 beta 1 integrins in all layers at intercellular junctions, and cytoplasmically in the lower strata. By contrast, alpha 6 beta 4 and occasionally alpha v beta 4 were expressed only by basal cells and localised to the basal lamina. These expression patterns were unaltered in specimens where an inflammatory cell infiltrate was present. In long-term cultures of normal urothelial cells maintained in a low-Ca++ serum-free medium, the monolayer cultures expressed alpha 2 beta 1, alpha 3 beta 1 and alpha 5 beta 1 integrins at intercellular junctions and in cytoplasmic inclusions, whereas alpha 6 beta 4 was distributed in a random pattern over the substratum. Increasing exogenous Ca++ concentrations induced cell stratification and desmosome formation, but not cytodifferentiation. Under these conditions, alpha 6 beta 4 became cell-, rather than substratum-associated, localising particularly to filopodia and lamellipodia. Quantitation of integrin expression by flow cytometry confirmed increased surface expression of alpha 6 beta 4 in high Ca++ media, and also of alpha 3 and alpha 5, but not alpha 2, subunits. These results suggest that alpha 2 beta 1 and alpha 3 beta 1 integrins, although differentially regulated, are mainly involved in homotypic cell-cell interactions and the maintenance of a stratified morphology, whereas alpha 6 beta 4 is the principal integrin involved in substratum adhesion.

Adolescent↗
PubMed1994

Interleukin-8 expression in Helicobacter pylori infected, normal, and neoplastic gastroduodenal mucosa.

AIMS: To investigate the expression of interleukin-8 (IL-8) in Helicobacter pylori infected normal and neoplastic gastroduodenal mucosa, and in established gastric cancer cell lines. METHODS: Immunofluorescence techniques were used to localise IL-8 in cryosections of gastric (n = 25) and duodenal (n = 17) endoscopic biopsy specimens an in resected gastric tumour tissue samples from 16 patients. Two gastric cancer cell lines (Kato 3 and MKN 45) were examined for IL-8 protein expression by immunofluorescence and for the presence of IL-8 mRNA by reverse transcription followed by the polymerase chain reaction (RT-PCR). RESULTS: IL-8 was localised to the epithelium in histologically normal gastric mucosa, with particularly strong expression in the surface cells. IL-8 expression was also a feature of surface epithelium in the duodenal bulb, but was much reduced in the second part of the duodenum. In chronic H pylori-associated gastritis gastritis gastric epithelial IL-8 expression was increased and expression of IL-8 within the lamina propria was evident. By contrast, large areas of IL-8 negative epithelium were observed in the body mucosa of a subject with Ménétrier's disease. In gastric carcinoma the tumour cells were positive for IL-8. IL-8 was also detected by immunofluorescence in unstimulated Kato 3 and MKN 45 cells, and constitutive IL-8 gene expression in these cell lines was confirmed by detection of IL-8 mRNA by RT-PCR. CONCLUSIONS: Immunoreactive IL-8, a potent neutrophil chemotactic and activating factor, is present in the epithelium of both normal and inflamed gastric mucosa with increased expression in the latter. There is site dependent variation in epithelial IL-8 expression within the gastroduodenal mucosa. The expression of the pro-inflammatory cytokine IL-8 in gastric carcinoma cells may influence peritumoural cellular infiltrates.

Base Sequence↗